To ascertain the safety of drugs under development, the pharmaceutical industry deploys a large amount of technology and process optimization to gain the ability of selecting and de-selecting drugs based on the criteria of therapeutic efficacy and potential of toxicity to the human health. This process is however not efficient, particularly because the potential of toxicity of drugs under development is not properly predicted by the current in vitro and in vivo models.
To date the evaluation of toxicity of a given substance relates to the use of animal models. Underlying is the assumption that toxic events observed in the animal are applicable and predictive of toxic events occurring in humans. The expression of animal toxicity data per dose at which toxicity is observed per body weight tends to undermine the occurrence of toxicity in humans. Moreover, the relatively low number of animals on which a substance is tested cannot predict for the occurrence of low frequency toxicity events that are shown in the human population. For example, a 0.01% incidence of toxicity in humans to a given substance means that approximately 25,000 to 250 million individuals are exposed to the given substance. To identify such a low occurrence toxicity in animals would require that 30 000 animals be exposed to the given substance. This being clearly impractical, exposure of fewer animals to high doses of a given substance is deemed desirable to identify hazards to human exposure to low doses.
The current drug development process only uses animal models to predict toxicity at a relatively late stage in drug development. The use of animal models would not be practical nor efficacious in generating toxicity data on the large number of substances found each year to have certain desirable therapeutic properties.
A significant need for alternative methods to predict toxicity exists.
Mechanistic approaches to model toxicity based on molecular mechanisms by which toxic event are expressed in humans have been pursued. Hence mitochondrial dysfunctions are studied in the context of pharmaco-toxicology using in vitro assays that describe one of the mitochondrial functions. These assays are used to measure acute, intrinsic toxicity of a given substance and are mostly based on the measure of oxidative phosphorylation in the several mitochondrial enzymatic complexes using either tissue biopsies, isolated mitochondria (Ishikawa et al., 2006), submitochondrial particles (Gustayson et al., 2002), or on the measure of cell ATP (Kerkela et al., 2006), or using cell viability assessment with the MTT test (Seok et al., 2007), or more recently on the measure of oxygen consumption using fluorescent or phosphorescent probes (Will et al., 2006).
The discovery that the mitochondrial membrane potential is essential to the normal functioning of mitochondria has led to the development of in vitro predictive assays to measure the effect of novel chemical entities on isolated mitochondria (Amacher, 2005). Similarly, observations that diseases affect mitochondrial motility has prompted the discovery of in vitro assays to identify substances that modify the changes in mobility induced by the disease in the hope of correcting the manifestations of the disease in humans (WO 2007/067809). However, these methods lack of predictivity and are hardly representative of the action of a tested substance on the entire mitochondria in its physiological context. Their use is limited to the screening of novel therapeutic substances or to the validation of a suspected mechanism of action of a therapeutic substance during its development.
Toxicity and lack of efficacy are currently the two main reasons for drug failure during development. Even though predictive toxicity data are acquired during the preclinical phases, 35 to 45% of the drug candidates fail due to unsuspected toxicity in humans. If some of the toxicity manifestations of a drug in development are seen during clinical trials, 20% of drugs among the 548 drugs approved by the food and drug administration (FDA) between 1975 and 1998, have caused toxic event of serious magnitude that motivated their change of label or market withdrawal. These facts raise the question of the efficacy of the current processes by which intrinsic, acute toxicity is assessed as a mean of predicting chronic or specific toxicities occurring in humans in the course of a therapeutic treatment.
Consequently, there is a strong need for early in vitro models that allow for a predictive assessment of the toxicity potential of substances, in particular for the ones with therapeutic properties during drug development.